SUMMARY Erythrocyte spectrin-actin junctions play a critical role in the maintenance of cell shape and membrane properties. Their organization and membrane linkage is therefore of fundamental importance to erythrocyte physiology. Our recent evidence indicates that dematin performs a pivotal function by linking these junctions to the membrane. Here, we propose that dematin performs its biological function by linking the spectrin-actin junctions to the membrane via glucose transporter-1 (GLUT1) in human erythrocytes. We will test this hypothesis under the following three aims: A1: GLUT1 interaction with the spectrin-actin junctions in human erythrocytes. We have identified GLUT1 as the primary membrane receptor for dematin and adducin in human erythrocytes. We propose to identify the interacting domains of GLUT1, dematin, and adducin, and disrupt the GLUT1-cytoskeletal bridge by biochemical means, thus assessing its impact on the membrane stability and shape of human erythrocytes. A2. Dematin and adducin binding receptor(s) in mouse erythrocytes and adipocytes. We propose that dematin and adducin bind to a novel membrane receptor(s) in mouse erythrocytes. We will identify this receptor using immunoprecipitation, mass spectrometry, and chemical crosslinking assays. We will test the binding of dematin and adducin with several potential membrane proteins identified in our proteomics screen of mouse erythrocyte membrane vesicles. We will also investigate the binding of dematin and adducin with GLUT4, and investigate their functional effects on GLUT4 recycling and glucose uptake in adipocytes. These experiments will clarify the role of alternate receptors that link the spectrin-actin junctions in mouse erythrocytes and adipocytes. A3. Physiological implications of complete dematin deficiency. Recently, we observed an obesity phenotype in the dematin headpiece domain null mice. To elucidate the specific role of the headpiece domain of dematin, we propose to generate a new mouse model with complete dematin deficiency using the conventional gene targeting techniques. We will compare the anemia and obesity phenotypes in the headpiece and dematin null mice focusing particularly on the impaired GLUT4 trafficking and diabetes. Together, these studies are likely to unveil a novel function of dematin, adducin, and glucose transporters in the regulation of anemia and metabolic disorders with broader implications for other headpiece domain-containing proteins.